The large calyx of Held nerve terminal is a pivotal element in the circuitry that computes sound source localization in the mammalian auditory brainstem. Precise timing of action potential output from this synapse is thought to be central for this task. However, the mechanisms that modulate and preserve action potential timing during high frequency firing are not well understood. The first hypothesis to be tested is that the efficiency of neurotransmitter (glutamate) release changes during development due to changes in release probability, which is controlled by the presynaptic action potential waveform and Ca buffering. We have found that action potential waveforms become faster and shorter in duration during the maturation of the calyx of Held. This will have important consequences for synaptic delays and release probability. We will use computer simulations and electrophysiology to determine how synaptic delays and the strength of synaptic communication varies with age. We will also test the hypothesis that the size of the readily releasable pool of synaptic vesicles increases during synapse maturation in order to compensate for a concomitant decrease in release probability. The second hypothesis is that diffusion plays the major role in fast glutamate clearance from the synaptic cleft of mature calyx synapses, whereas metabotropic glutamate receptors (mGluRs) in immature synapses act as autoreceptors that limit the amount of glutamate release. Too much glutamate can be toxic for the brain, and it may also disrupt synaptic transmission by desensitizing ionotropic receptors, so we propose that mGluRs may play a neuroprotective role early during development, limiting glutamate release at a time when glia are still immature. The third hypothesis is that the Na+/K+ ATPase plays a major role in the ability of the calyx of Held to fire action potentials at high frequencies. We will determine the location of different subtypes of the Na+/K+-ATPases and how their function affects presynaptic and postsynaptic physiology. The auditory system consumes the highest amount of glucose in the human brain. Most of that energy in the brain is consumed by the Na+/K+-ATPase as it restores and maintains the ionic gradients of Na+ and K* ions across the plasma membrane. Presumably due to its unusually high rate of spiking, the auditory brainstem and cortex consume remarkably large amounts of ATP, principally via the operation of the Na+/K+-ATPase. However, the properties of presynaptic Na+/K*-ATPases are unknown due to the small size of most CMS nerve terminals. Little is also known about the location, subtype and functional properties of Na+/K+-ATPases in auditory pathways. By blocking Na*/K+-ATPases with specific pharmacological agents we will mimic the effects of ischemia and strokes, when neurons are starved for oxygen and glucose. Thus, this research proposal will generate basic data and insights on a critical component of energy consumption in the brain.